New beta-tubulin compositions, methods of production and uses of beta-tubulin compositions

Abstract

The present invention provides a new heat-treated beta-casein composition characterized by a low molar ratio between the combined content of calcium and magnesium and the combined content of beta-casein and gamma-casein. The present invention also provides a method for preparing such heat-treated beta-casein composition and the use of such heat-treated beta-casein composition.

Description

Invention Field

[0001] This invention provides novel heat-treated β-casein compositions characterized by a low molar ratio between the combined content of calcium and magnesium and the combined content of β-casein and γ-casein. The invention also provides a method for preparing such heat-treated β-casein compositions and uses of such heat-treated β-casein compositions. background

[0002] From a nutritional perspective, β-casein is an interesting milk protein and is valuable, for example, in the production of infant formula products. β-casein exists in mammalian milk along with other proteins such as α-casein, κ-casein, and lactose albumin such as β-lactoglobulin, α-lactalbumin, and serum albumin.

[0003] Several methods for isolating β-casein have been proposed in this field: WO 2012 / 148,269 A1 discloses a method for preparing a milk protein fraction containing β-casein. The method involves subjecting milk to a first microfiltration step, and then subjecting the residue from the first microfiltration step to a second microfiltration step. The first microfiltration step can be performed at a warm temperature, and the second microfiltration step can be performed at a low temperature.

[0004] US 2007 / 0104847 A1 discloses a method for isolating β-casein from milk. This method is based on initial cold microfiltration of cooled skim milk to obtain a partially β-casein-depleted permeate and a permeate containing milk serum proteins and a significant amount of β-casein. The permeate containing β-casein can undergo further purification.

[0005] FR 2,592,769 A discloses the production of β-casein by microfiltration of a cooled liquid feed containing aggregated calcium caseinate.

[0006] US 5,169,666 A describes a process for producing a milk protein product rich in β-casein by microfiltration of cooled skim milk. The microfiltration membrane used has a pore size of 0.1 micrometer to 0.2 micrometer. Invention Overview

[0007] The inventors have discovered that protein solutions containing high concentrations of β-casein are difficult to process and handle, and have observed sudden clogging of processing equipment and the development of undesirable viscosity during and after heat treatment. The inventors have also observed indications that the use of β-casein may lead to undesirable sensory changes in the final product, for example, in experimental high-protein beverages based on β-casein as the primary protein source, and may cause processing problems during the production of β-casein-rich infant formulas.

[0008] The inventors have investigated these problems and have surprisingly discovered that they can be reduced or even avoided by controlling the molar ratio between the combined content of calcium and magnesium and the combined content of β-casein and γ-casein. The heat-treated β-casein concentrate of this invention appears less prone to undesirable thickening during the production of β-casein-rich foods, as well as during subsequent storage and use of these foods.

[0009] Therefore, aspects of the present invention relate to a heat-treated β-casein concentrate having: The total content of bCN (bCN) and γ-casein (gCN) relative to the weight of hbc is at least 9% w / w. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein. The molar ratio between the following items is at most 6.0: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

[0010] Another aspect of the present invention relates to a method for producing the heat-treated β-casein concentrate of the present invention, the method comprising the following steps: a) Provide a protein solution (ps), said protein solution (ps) having: The total content of bCN and γ-casein (gCN) relative to the weight of the protein solution is at least 9% w / w. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein. Preferably, the molar ratio between at most 6.0 of the following items is: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN, and pH range of 6-9 b) Heat the protein solution from step a) to a temperature of at least 65°C, and c) Optionally, the heat-treated protein solution or its protein concentrate is dried.

[0011] Another aspect of the invention relates to a process for producing a food containing bCN and optionally also containing gCN or its protein hydrolysates, the method comprising the following steps: I) Provides the heat-treated bCN concentrate of the present invention. II) subjecting the heat-treated bCN concentrate to further processing, such as involving the hydrolysis of bCN and / or mixing with other components and / or filling the heat-treated bCN concentrate into a suitable container.

[0012] Another aspect of the present invention relates to a food product obtainable by the process of the present invention.

[0013] Another aspect of the invention relates to the use of the heat-treated bCN concentrate of the invention as a food ingredient, preferably for the production of one or more of the foods mentioned herein, preferably using one or more features of the processes mentioned herein. Detailed Explanation

[0014] An aspect of the present invention relates to a heat-treated β-casein concentrate having: The total content of β-casein (bCN) and γ-casein (gCN) relative to the weight of hbc is at least 9% w / w. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein. The molar ratio between the following items is at most 6.0: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

[0015] Preferably, the heat-treated β-casein concentrate has: The total content of bCN and gCN is at least 9% w / w relative to the weight of hbc. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein. The molar ratio between the following items is at most 6.0: The total content of calcium and magnesium, and The total content of bCN and gCN.

[0016] In the context of this invention, the term "β-casein" (abbreviated as bCN) has its general meaning and covers naturally occurring variants of β-casein present in mammals. The term also covers proteins having the amino acid sequence of β-casein but which have been prepared, for example, by recombinant expression or by fermentation. Further details regarding bovine β-casein can be found, for example, in McSweeney ("Advanced Dairy Chemistry, Volume 1: Proteins"; 3rd ed., Kluwer Academic Publishers, 2003; Chapter 3) and in... https: / / www.uniprot.org / uniprotkb / P02666 / entryIt was found in bovine β-casein, and more details about human β-casein can be found in [website / source]. https: / / www.uniprot.org / uniprotkb / W5RWE1 / entry It was found in (human β-casein).

[0017] In the context of this invention, the term "γ-casein" (abbreviated as gCN) has its general meaning. γ-casein is a hydrophobic peptide produced by plasmin-based proteolytic hydrolysis of β-casein. Three distinct γ-caseins (γ-1-casein, γ-2-casein, and γ-3-casein) are typically found, corresponding to three different cleavage sites in β-casein. The term "γ-casein" encompasses naturally occurring variants of γ-casein. The term also encompasses proteins having the amino acid sequence of γ-casein but which have been prepared, for example, by fermentation. Further details about γ-casein can be found at McSweeney.

[0018] Based on the analysis A mentioned in this article, the combined content of bCN and gCN, as well as the relative content of bCN and gCN alone, were determined.

[0019] In the context of this invention, the term "heat-treated β-casein concentrate" (abbreviated as "hbc") refers to the β-casein product of this invention and may be, for example, in liquid or powder form. hbc contains at least 50% w / w of β-casein relative to the total protein. Preferably, hbc contains at least 25% w / w of β-casein relative to the total solids. The term "heat-treated" with respect to hbc means that the preparation of hbc involves a heat treatment step, wherein the liquid bCN concentrate has been heated to a temperature of at least 65°C, preferably by heating the liquid bCN concentrate to at least 65°C to reduce the microbial content and / or by subjecting the liquid bCN concentrate to a drying step, such as, for example, spray drying, which involves heat treatment to at least 65°C. Therefore, hbc typically has a low microbial content. hbc is suitable for use as a food ingredient and is therefore edible. hbc is preferably suitable for use as a powdered food ingredient for the production of infant formula by dry blending.

[0020] In the context of this invention, the term "molar ratio" or "weight ratio" between entity A and entity B refers to ratio n. A / n B or w A / w B And it is expressed as a decimal number. For example, if the composition contains 9% by weight of entity A and 3% by weight of entity B, then the weight ratio of A to B in the composition is 9% / 3% = 3. Similarly, if the composition contains 4 moles of entity A and 2 moles of entity B, then the molar ratio of A to B in the composition is 4 / 2 = 2.

[0021] In some preferred embodiments of the invention, the protein of HBC is a mammalian milk protein, such as human milk protein or optionally a ruminant milk protein. Particularly preferred are the milk proteins of one or more of the following: cow, sheep, goat, buffalo, camel, llama, mare, and deer. Even more preferred are the milk proteins of HBC that are present in cow's milk.

[0022] In other preferred embodiments of the invention, the bCN and gCN of hbc have amino acid sequences of mammalian bCN and gCN, such as human bCN and gCN, or optionally ruminant bCN and gCN. Particularly preferred are the bCN and gCN of hbc having amino acid sequences of bCN and gCN found in one or more of cattle, sheep, goats, buffalo, camels, llamas, mares, and deer. Even more preferred are the bCN and gCN of hbc having amino acid sequences of bCN and gCN found in cow's milk.

[0023] In some preferred embodiments of the invention, the heat-treated bCN concentrate is in powder form, preferably having a total solids content of at least 90% w / w, more preferably at least 92% w / w, even more preferably at least 94% w / w, and most preferably at least 95% w / w.

[0024] In other preferred embodiments of the invention, the heat-treated bCN concentrate is in liquid form, preferably having a total solids content in the range of 9% w / w-50% w / w, more preferably 9% w / w-40% w / w, even more preferably 10% w / w-30% w / w, and most preferably 11% w / w-25% w / w.

[0025] The heat-treated bCN concentrate in liquid form is preferably an aqueous solution, and water preferably constitutes the majority (if not all) of the non-solid matter in the heat-treated bCN concentrate in liquid form. In some preferred embodiments of the invention, water contributes at least 90% w / w, more preferably at least 95% w / w, and most preferably about 100% w / w of the non-solid matter in the heat-treated bCN concentrate in liquid form. The content of the non-solid matter in the composition is determined as 100% minus the total solids content.

[0026] In some preferred embodiments of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is at least 9% w / w relative to the weight of hbc, more preferably at least 9.5% w / w, even more preferably at least 10% w / w, and most preferably at least 11% w / w relative to the weight of hbc.

[0027] In some preferred embodiments of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is 9% w / w-25% w / w, more preferably 9% w / w-20% w / w, even more preferably 9.5% w / w-16% w / w, and most preferably 10% w / w-14% w / w relative to the weight of hbc.

[0028] In other preferred embodiments of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is 9% w / w-14% w / w, more preferably 9% w / w-13% w / w, even more preferably 9.5% w / w-12% w / w, and most preferably 9.5% w / w-11% w / w relative to the weight of hbc.

[0029] In some preferred embodiments of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is at least 40% w / w relative to the weight of hbc, more preferably at least 50% w / w, even more preferably at least 60% w / w, and most preferably at least 70% w / w relative to the weight of hbc.

[0030] In other preferred embodiments of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is at least 80% w / w relative to the weight of hbc, more preferably at least 90% w / w, even more preferably at least 95% w / w, and most preferably at least 98% w / w relative to the weight of hbc.

[0031] In some preferred embodiments of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is 40% w / w-98% w / w, more preferably 50% w / w-95% w / w, even more preferably 55% w / w-90% w / w, and most preferably 60% w / w-85% w / w relative to the weight of hbc.

[0032] Preferably, the heat-treated bCN concentrate has a molar ratio of at most 1.0 between gCN and bCN.

[0033] In the context of this invention, the terms "molar ratio" for gCN and bCN refer to a value obtained through calculation (m gCN / m bCN ), where m gCN It is the content of gCN in moles, and m bCN This refers to the content of bCN in moles.

[0034] In other preferred embodiments of the invention, the molar ratio between gCN and bCN in the heat-treated bCN concentrate is at most 0.67, more preferably at most 0.50, even more preferably at most 0.25, and most preferably at most 0.15.

[0035] In some preferred embodiments of the present invention, the molar ratio between gCN and bCN in the heat-treated bCN concentrate is 0-1.0, more preferably 0.001-0.67, more preferably 0.005-0.50, even more preferably 0.01-0.25, and most preferably 0.02-0.15.

[0036] In other preferred embodiments of the invention, the molar ratio between gCN and bCN in the heat-treated bCN concentrate is 0.05-1.0, more preferably 0.1-1.0, even more preferably 0.2-1.0, and most preferably 0.3-1.0.

[0037] In another preferred embodiment of the invention, the molar ratio between gCN and bCN in the heat-treated bCN concentrate is 0.02-1.0, more preferably 0.05-0.6, and most preferably 0.08-0.4.

[0038] In some preferred embodiments of the invention, the heat-treated bCN concentrate does not contain gCN.

[0039] Generally, it is preferred that the β-casein and γ-casein of the heat-treated bCN concentrate have been isolated from mammalian milk or milk serum, preferably from ruminant milk, and more preferably from cow's milk.

[0040] Furthermore, it is generally preferred that the β-casein and γ-casein in the heat-treated bCN concentrate have the same degree of phosphorylation as they would have in mammalian milk. The degree of phosphorylation can be determined by LC-MS.

[0041] In addition, the phosphorylation level of β-casein and / or γ-casein can be altered by subjecting the protein to a phosphorylation step to increase the degree of phosphorylation or by subjecting the protein to a dephosphorylation step to decrease the degree of phosphorylation.

[0042] However, in other preferred embodiments of the invention, the β-casein and γ-casein of the heat-treated bCN concentrate are not phosphorylated. This is, for example, if the β-casein and γ-casein (if the latter is present) are prepared by fermentation in a host organism or under conditions that do not produce phosphorylation during post-translational modification, or if the protein has undergone a dephosphorylation step.

[0043] In some preferred embodiments of the invention, the heat-treated bCN concentrate contains a total protein content of at least 50% w / w, more preferably at least 70% w / w, even more preferably at least 80% w / w, and most preferably at least 90% w / w relative to the total solids.

[0044] Total protein and total solids were measured according to Examples 1.5 and 1.15 of WO2020 / 002450 A1.

[0045] In another preferred embodiment of the invention, the heat-treated bCN concentrate contains a total protein content in the range of 6% w / w-50% w / w, more preferably 8% w / w-40% w / w, even more preferably 10% w / w-30% w / w, and most preferably 12% w / w-20% w / w.

[0046] In some preferred embodiments of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is at least 25% w / w relative to the total protein, and more preferably at least 40% w / w relative to the total protein.

[0047] In another preferred embodiment of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is at least 50% w / w relative to the total protein, more preferably at least 60% w / w, even more preferably at least 70% w / w, and most preferably at least 80% w / w relative to the total protein.

[0048] In even more preferred embodiments of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is at least 85% w / w relative to the total protein content, more preferably at least 90% w / w, and most preferably at least 95% w / w relative to the total protein content.

[0049] In some preferred embodiments of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is 40% w / w-98% w / w, more preferably 50% w / w-96% w / w, even more preferably 60% w / w-94% w / w, and most preferably 70% w / w-92% w / w relative to the total protein.

[0050] In other preferred embodiments of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is 40% w / w-80% w / w, more preferably 40% w / w-75% w / w, even more preferably 40% w / w-70% w / w, and most preferably 40% w / w-65% w / w relative to the total protein.

[0051] In some particularly preferred embodiments of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is at least 70% w / w, more preferably at least 80% w / w, even more preferably at least 90% w / w, even more preferably at least 95% w / w, and most preferably at least 98% w / w relative to the total amount of casein.

[0052] The total amount of casein was determined according to ISO 17997-1:2004.

[0053] In another preferred embodiment of the invention, the total content of bCN and gCN in the heat-treated bCN concentrate is 65% w / w-98% w / w, more preferably 70% w / w-96% w / w, even more preferably 75% w / w-94% w / w, and most preferably 80% w / w-94% w / w relative to the total amount of casein.

[0054] In some preferred embodiments of the invention, the heat-treated bCN concentrate contains whey protein in an amount of 10% w / w to 40% w / w, more preferably 15% w / w to 35% w / w, and most preferably 20% w / w to 30% w / w, relative to non-bCN protein.

[0055] In some preferred embodiments of the invention, the heat-treated bCN concentrate contains whey protein in an amount of at least 50% w / w relative to non-bCN protein, more preferably at least 70% w / w, and most preferably at least 90% w / w relative to non-bCN protein.

[0056] In the context of this invention, the term "non-bCN protein" refers to proteins that are not bCN and includes, for example, γ-casein, whey protein, peptides that contribute to total protein, and other protein substances that may be present.

[0057] In some preferred embodiments of the invention, the heat-treated bCN concentrate contains 60% w / w-90% w / w, more preferably 65% ​​w / w-85% w / w, and most preferably 70% w / w-80% w / w, relative to the non-bCN protein.

[0058] The term "bCN peptide" refers to peptides that are obtained or available through partial proteolytic hydrolysis of bCN, and includes γ-casein.

[0059] In some preferred embodiments of the invention, the heat-treated bCN concentrate has a molar ratio between at most 5, more preferably at most 4, even more preferably at most 3, and most preferably at most 2.5 of the following: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

[0060] In another preferred embodiment of the invention, the heat-treated bCN concentrate has a molar ratio between at most 2.0, more preferably at most 1.6, even more preferably at most 1.3, and most preferably at most 1.0: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

[0061] In some preferred embodiments of the invention, the heat-treated bCN concentrate has a molar ratio between at most 6, more preferably at most 5, even more preferably at most 4, even more preferably at most 3, and most preferably at most 2.5 of the following: The total content of calcium and magnesium, and The total content of bCN and gCN.

[0062] In another preferred embodiment of the invention, the heat-treated bCN concentrate has a molar ratio between at most 2.0, more preferably at most 1.6, even more preferably at most 1.3, and most preferably at most 1.0: The total content of calcium and magnesium, and The total content of bCN and gCN.

[0063] Other divalent metal cations, such as Zn, for example 2+ or Fe 2+ It may also be present in heat-treated bCN concentrates, but typically in amounts significantly lower than those of calcium and magnesium. In some preferred embodiments of the invention, the total weight of the divalent metal cations of hbc other than calcium and magnesium is at most 50%, more preferably at most 25%, and even more preferably at most 10% of the total weight of calcium and magnesium.

[0064] HBC typically contains monovalent metal ions, such as, for example, sodium and potassium. In some preferred embodiments of the invention, the total amount of monovalent metal cations contained in HBC is in the range of 0.1% w / w to 2% w / w relative to the total protein, more preferably 0.13% w / w to 1.8% w / w relative to the total protein, and most preferably 0.15% w / w to 1.8% w / w relative to the total protein.

[0065] In other preferred embodiments of the invention, the total amount of monovalent metal cations contained in HBC is in the range of 0.5% w / w to 1.8% w / w relative to the total protein, more preferably 0.6% w / w to 1.4% w / w relative to the total protein. These ranges are often present, for example, when cation exchange is used to adjust the calcium and magnesium content.

[0066] In some preferred embodiments of the invention, the heat-treated bCN concentrate contains at most 15% w / w, more preferably at most 10% w / w, even more preferably at most 5% w / w, and most preferably at most 1% w / w.

[0067] In some preferred embodiments of the invention, the heat-treated bCN concentrate contains carbohydrates in an amount of up to 50% w / w relative to total solids, more preferably up to 40% w / w relative to total solids, even more preferably up to 30% w / w relative to total solids, and most preferably up to 20% w / w relative to total solids.

[0068] In another preferred embodiment of the invention, the heat-treated bCN concentrate contains carbohydrates in an amount of up to 15% w / w relative to total solids, more preferably up to 10% w / w relative to total solids, even more preferably up to 5% w / w relative to total solids, and most preferably up to 1% w / w relative to total solids.

[0069] In some preferred embodiments of the invention, the heat-treated bCN concentrate contains at most 15% w / w, more preferably at most 10% w / w, even more preferably at most 5% w / w, and most preferably at most 1% w / w.

[0070] In some preferred embodiments of the invention, the heat-treated bCN concentrate contains lactose in an amount of up to 50% w / w relative to total solids, more preferably up to 40% w / w relative to total solids, even more preferably up to 30% w / w relative to total solids, and most preferably up to 20% w / w relative to total solids.

[0071] In another preferred embodiment of the invention, the heat-treated bCN concentrate contains lactose in an amount of up to 15% w / w relative to total solids, more preferably up to 10% w / w relative to total solids, even more preferably up to 5% w / w relative to total solids, and most preferably up to 1% w / w relative to total solids.

[0072] In some preferred embodiments of the invention, the heat-treated bCN concentrate contains at most 5% w / w, more preferably at most 2% w / w, even more preferably at most 1% w / w, and most preferably at most 0.2% w / w.

[0073] In other preferred embodiments of the invention, the heat-treated bCN concentrate contains lipids in an amount of up to 15% w / w relative to total solids, more preferably up to 12% w / w relative to total solids, even more preferably up to 10% w / w relative to total solids, and most preferably up to 8% w / w relative to total solids.

[0074] In another preferred embodiment of the invention, the heat-treated bCN concentrate contains lipids in an amount of up to 5% w / w relative to total solids, more preferably up to 2% w / w relative to total solids, even more preferably up to 1% w / w relative to total solids, and most preferably up to 0.2% w / w relative to total solids. This type of heat-treated bCN concentrate is particularly preferred for low-fat applications or applications using other lipid sources.

[0075] In some preferred embodiments of the invention, the pH of the heat-treated bCN concentrate is in the range of 6-9, more preferably 6.1-8.0, even more preferably 6.3-7.5, and most preferably 6.5-7.2.

[0076] pH was measured according to Example 1.16 of WO2020 / 002450 A1. The pH values ​​mentioned herein refer to pH normalized to 25°C.

[0077] In other preferred embodiments of the invention, the pH of the heat-treated bCN concentrate is in the range of 6.5-9, more preferably 6.7-8.0, even more preferably 7.0-8.0, and most preferably 7.1-8.0.

[0078] In some preferred embodiments of the present invention, the heat-treated bCN concentrate has: pH in the range of 6-7.0, and The molar ratios between at most 5, more preferably at most 4, even more preferably at most 3, and most preferably at most 2.5 of the following: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

[0079] In other preferred embodiments of the invention, the heat-treated bCN concentrate has: pH in the range of 6-7.0, and The molar ratio between at most 2.0, more preferably at most 1.6, even more preferably at most 1.3, and most preferably at most 1.0 is: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

[0080] In some preferred embodiments of the present invention, the heat-treated bCN concentrate has: pH in the range of 6-7.0, and The molar ratios between at most 5, more preferably at most 4, even more preferably at most 3, and most preferably at most 2.5 of the following: The total content of calcium and magnesium, and The total content of bCN and gCN.

[0081] In other preferred embodiments of the invention, the heat-treated bCN concentrate has: pH in the range of 6-7.0, and The molar ratio between at most 2.0, more preferably at most 1.6, even more preferably at most 1.3, and most preferably at most 1.0 is: The total content of calcium and magnesium, and The total content of bCN and gCN.

[0082] The advantage of the heat-treated bCN concentrate of the present invention is that it can be obtained with a low content of colony-forming units, making it suitable for the production of infant formula. The powder variant of the heat-treated bCN concentrate of the present invention is also suitable for preparing infant formula products by dry blending, i.e., by blending the powder components to obtain infant formula in powder form.

[0083] In some preferred embodiments of the present invention, the colony forming unit (CFU) content of the heat-treated bCN concentrate is up to 10,000 CFU / g total solids, more preferably up to 5,000 CFU / g total solids, even more preferably up to 1,000 CFU / g total solids, and most preferably up to 100 CFU / g total solids.

[0084] Preferably, the heat-treated bCN concentrate is sterile. Therefore, in some preferred embodiments of the invention, the heat-treated bCN concentrate has been heat-sterilized.

[0085] Another advantage of the heat-treated bCN concentrate of the present invention in powder form is that it can be obtained with a relatively high bulk density, which makes it suitable for producing powdered foods, such as infant formula, by dry blending and reduces the risk of powder particle separation in dry blended foods.

[0086] In some preferred embodiments of the invention, the heat-treated bCN concentrate is in powder form, preferably prepared by spray drying, and has a bulk density of at least 0.20 g / cm³. 3 More preferably at least 0.22 g / cm³ 3 And most preferably at least 0.24 g / cm³ 3 .

[0087] The bulk density was measured according to Example 1.17 of WO2020 / 002450 A1.

[0088] In another preferred embodiment of the invention, the heat-treated bCN concentrate is in powder form, the powder having a bulk density of 0.20 g / cm³. 3 -1.0 g / cm 3 More preferably 0.21 g / cm 3 -0.6 g / cm 3 And the most preferred value is 0.22 g / cm³. 3 -0.4 g / cm 3 Within the range.

[0089] The advantage of the heat-treated bCN concentrate of the present invention is that it exhibits relatively low viscosity even when present in solution at relatively high concentrations. This makes the heat-treated bCN concentrate suitable for high-protein applications, such as bCN-based high-protein beverages or for processing using high concentrations of bCN.

[0090] In some preferred embodiments of the invention, the viscosity of the heat-treated bCN concentrate is such that, when adjusted to a protein content of 10.0% w / w, it is obtained at 25°C and 145 s. -1 The shear rate is at most 30 mPa*s, more preferably at 145 s at 25°C. -1 The shear rate is at most 25 mPa*s, and even more preferably at 25°C for 145 s. -1 The shear rate is at most 22 mPa*s, and most preferably at 25°C for 145 s. -1 The shear rate is at most 20 mPa*s.

[0091] If the heat-treated bCN concentrate is in powder form, the viscosity is measured by dissolving the powder in distilled water to obtain a protein content of 10.0% w / w.

[0092] Viscosity was measured using a rheometer method according to Example 1.8 of WO2020 / 002450 A1.

[0093] In some particularly preferred embodiments of the invention, the heat-treated β-casein concentrate is in powder form and has the following properties: The total content of bCN and gCN is at least 40% w / w relative to the weight of hbc, and more preferably at least 60% w / w relative to the weight of hbc. The molar ratio between gCN and bCN is at most 1.0, more preferably at most 0.67. pH in the range of 6.3-7.5, more preferably 6.5-7.2. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein, and more preferably at least 90% w / w relative to the total amount of casein. The molar ratio between at most 6.0, more preferably at most 2, of the following items: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

[0094] In other particularly preferred embodiments of the invention, the heat-treated β-casein concentrate is in powder form and has the following properties: The total content of bCN and gCN is at least 40% w / w relative to the weight of hbc, and more preferably at least 60% w / w relative to the weight of hbc. The molar ratio between gCN and bCN is at most 1.0, more preferably at most 0.67. pH in the range of 6.3-7.5, more preferably 6.5-7.2. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein, and more preferably at least 90% w / w relative to the total amount of casein. The molar ratio between the following items is at most 6.0, more preferably at most 2.0: The total content of calcium and magnesium, and The total content of bCN and gCN.

[0095] In some particularly preferred embodiments of the invention, the heat-treated β-casein concentrate is in powder form and has the following properties: The total content of bCN and gCN is at least 40% w / w relative to the weight of hbc, and more preferably at least 60% w / w relative to the weight of hbc. The molar ratio between gCN and bCN is at most 1.0, more preferably at most 0.67. pH in the range of 6.3-7.5, more preferably 6.5-7.2. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein, and more preferably at least 90% w / w relative to the total amount of casein. The molar ratio between at most 6.0, more preferably at most 2, of the following items: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN, and At least 0.20 g / cm 3 More preferably at least 0.22 g / cm³ 3 The packing density within the range.

[0096] In other particularly preferred embodiments of the invention, the heat-treated β-casein concentrate is in powder form and has the following properties: The total content of bCN and gCN is at least 40% w / w relative to the weight of hbc, and more preferably at least 60% w / w relative to the weight of hbc. The molar ratio between gCN and bCN is at most 1.0, more preferably at most 0.67. pH in the range of 6.3-7.5, more preferably 6.5-7.2. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein, and more preferably at least 90% w / w relative to the total amount of casein. The molar ratio between the following items is at most 6.0, more preferably at most 2.0: The total content of calcium and magnesium, and The total content of bCN and gCN, and At least 0.20 g / cm 3 More preferably at least 0.22 g / cm³ 3 The packing density within the range.

[0097] In some particularly preferred embodiments of the invention, the heat-treated β-casein concentrate is in powder form and has the following properties: The total content of bCN and gCN is at least 40% w / w relative to the weight of hbc, and more preferably at least 60% w / w relative to the weight of hbc. The molar ratio between gCN and bCN is at most 1.0, more preferably at most 0.67. pH in the range of 6.3-7.5, more preferably 6.5-7.2. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein, and more preferably at least 90% w / w relative to the total amount of casein. The molar ratio between at most 6.0, more preferably at most 2, of the following items: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN, and The content of colony-forming units is up to 10,000 CFU / g total solids, more preferably up to 1,000 CFU / g total solids.

[0098] In other particularly preferred embodiments of the invention, the heat-treated β-casein concentrate is in powder form and has the following properties: The total content of bCN and gCN is at least 40% w / w relative to the weight of hbc, and more preferably at least 60% w / w relative to the weight of hbc. The molar ratio between gCN and bCN is at most 1.0, more preferably at most 0.67. pH in the range of 6.3-7.5, more preferably 6.5-7.2. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein, and more preferably at least 90% w / w relative to the total amount of casein. The molar ratio between the following items is at most 6.0, more preferably at most 2.0: The total content of calcium and magnesium, and The total content of bCN and gCN, and The content of colony-forming units is up to 10,000 CFU / g total solids, more preferably up to 1,000 CFU / g total solids.

[0099] In some particularly preferred embodiments of the invention, the heat-treated β-casein concentrate is in powder form and has the following properties: The total content of bCN and gCN is at least 40% w / w relative to the weight of hbc, and more preferably at least 60% w / w relative to the weight of hbc. The molar ratio between gCN and bCN is at most 1.0, more preferably at most 0.67. pH in the range of 6.3-7.5, more preferably 6.5-7.2. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein, and more preferably at least 90% w / w relative to the total amount of casein. The molar ratio between at most 6.0, more preferably at most 2, of the following items: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN At least 0.20 g / cm 3 More preferably at least 0.22 g / cm³ 3 The packing density within the range, and The content of colony-forming units is up to 10,000 CFU / g total solids, more preferably up to 1,000 CFU / g total solids.

[0100] In other particularly preferred embodiments of the invention, the heat-treated β-casein concentrate is in powder form and has the following properties: The total content of bCN and gCN is at least 40% w / w relative to the weight of hbc, and more preferably at least 60% w / w relative to the weight of hbc. The molar ratio between gCN and bCN is at most 1.0, more preferably at most 0.67. pH in the range of 6.3-7.5, more preferably 6.5-7.2. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein, and more preferably at least 90% w / w relative to the total amount of casein. The molar ratio between the following items is at most 6.0, more preferably at most 2.0: The total content of calcium and magnesium, and The total content of bCN and gCN, and At least 0.20 g / cm 3 More preferably at least 0.22 g / cm³ 3 The packing density within the range, and The content of colony-forming units is up to 10,000 CFU / g total solids, more preferably up to 1,000 CFU / g total solids.

[0101] In some particularly preferred embodiments of the invention, the heat-treated β-casein concentrate is in powder form and has the following properties: The total content of bCN and gCN is at least 40% w / w relative to the weight of hbc, and more preferably at least 60% w / w relative to the weight of hbc. The molar ratio between gCN and bCN is at most 1.0, more preferably at most 0.67. pH in the range of 6.3-7.5, more preferably 6.5-7.2. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein, and more preferably at least 90% w / w relative to the total amount of casein. The molar ratio between at most 6.0, more preferably at most 2, of the following items: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN, and When adjusted to a protein content of 10.0% w / w at 25°C and 145 s -1 The shear rate is at most 30 mPa*s, more preferably at 145 s at 25°C. -1 The viscosity has a shear rate of up to 25 mPa*s.

[0102] In other particularly preferred embodiments of the invention, the heat-treated β-casein concentrate is in powder form and has the following properties: The total content of bCN and gCN is at least 40% w / w relative to the weight of hbc, and more preferably at least 60% w / w relative to the weight of hbc. The molar ratio between gCN and bCN is at most 1.0, more preferably at most 0.67. pH in the range of 6.3-7.5, more preferably 6.5-7.2. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein, and more preferably at least 90% w / w relative to the total amount of casein. The molar ratio between the following items is at most 6.0, more preferably at most 2.0: The total content of calcium and magnesium, and The total content of bCN and gCN, and When adjusted to a protein content of 10.0% w / w at 25°C and 145 s -1 The shear rate is at most 30 mPa*s, more preferably at 145 s at 25°C. -1 The viscosity has a shear rate of up to 25 mPa*s.

[0103] In some particularly preferred embodiments of the invention, the heat-treated β-casein concentrate is in powder form and has the following properties: The total content of bCN and gCN is at least 40% w / w relative to the weight of hbc, and more preferably at least 60% w / w relative to the weight of hbc. The molar ratio between gCN and bCN is at most 1.0, more preferably at most 0.67. pH in the range of 6.3-7.5, more preferably 6.5-7.2. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein, and more preferably at least 90% w / w relative to the total amount of casein. The molar ratio between at most 6.0, more preferably at most 2, of the following items: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN The content of colony-forming units is up to 10,000 CFU / g total solids, more preferably up to 1,000 CFU / g total solids, and When adjusted to a protein content of 10.0% w / w at 25°C and 145 s -1 The shear rate is at most 30 mPa*s, more preferably at 145 s at 25°C. -1 The viscosity has a shear rate of up to 25 mPa*s.

[0104] In other particularly preferred embodiments of the invention, the heat-treated β-casein concentrate is in powder form and has the following properties: The total content of bCN and gCN is at least 40% w / w relative to the weight of hbc, and more preferably at least 60% w / w relative to the weight of hbc. The molar ratio between gCN and bCN is at most 1.0, more preferably at most 0.67. pH in the range of 6.3-7.5, more preferably 6.5-7.2. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein, and more preferably at least 90% w / w relative to the total amount of casein. The molar ratio between the following items is at most 6.0, more preferably at most 2.0: The total content of calcium and magnesium, and The total content of bCN and gCN The content of colony-forming units is up to 10,000 CFU / g total solids, more preferably up to 1,000 CFU / g total solids, and When adjusted to a protein content of 10.0% w / w at 25°C and 145 s -1 The shear rate is at most 30 mPa*s, more preferably at 145 s at 25°C. -1 The viscosity has a shear rate of up to 25 mPa*s.

[0105] Another aspect of the present invention relates to a method for producing the heat-treated β-casein concentrate (hbc) of the present invention, the method comprising the following steps: a) Provide a protein solution (ps), said protein solution (ps) having: The total content of β-casein (bCN) and γ-casein (gCN) relative to at least 9% w / w of the protein solution weight. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein. Preferably, the molar ratio between at most 6.0 of the following items is: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN, and pH range of 6-9 b) Heat the protein solution from step a) to a temperature of at least 65°C, and c) Optionally, the heat-treated protein solution or its protein concentrate is dried.

[0106] In the context of this invention, the term "dry" means that the composition or product in question contains up to 10% (w / w) water, and preferably even less.

[0107] Step a) involves providing a protein solution, which is heat-treated in step b). The protein solution is an aqueous solution, and water preferably constitutes the majority (if not all) of the non-solid matter of the protein solution. In some preferred embodiments of the invention, water contributes at least 90% w / w, more preferably at least 95% w / w, and most preferably about 100% w / w of the non-solid matter of the protein solution. The content of the non-solid matter in the composition is determined as 100% minus the total solids content.

[0108] In some preferred embodiments of the invention, the total content of bCN and gCN in the protein solution of step a) of the method for producing hbc is at least 9% w / w relative to the weight of ps, more preferably at least 9.5% w / w, even more preferably at least 10% w / w, and most preferably at least 11% w / w relative to the weight of ps.

[0109] In another preferred embodiment of the invention, the total content of bCN and gCN in the protein solution of step a) is 9% w / w-25% w / w, more preferably 9% w / w-20% w / w, even more preferably 9.5% w / w-16% w / w, and most preferably 10% w / w-14% w / w relative to the weight of ps.

[0110] In other preferred embodiments of the invention, the total content of bCN and gCN in the protein solution of step a) is 9% w / w-14% w / w, more preferably 9% w / w-13% w / w, even more preferably 9.5% w / w-12% w / w, and most preferably 9.5% w / w-11% w / w relative to the weight of ps.

[0111] Except for the characteristics and preferences that apply only to HBC in powder form, the characteristics and preferences described for HBC also apply to PS.

[0112] In some preferred embodiments of the invention, the protein solution of step a) has a molar ratio of at most 1.0 between gCN and bCN.

[0113] In other preferred embodiments of the invention, the molar ratio between gCN and bCN in the protein solution of step a) is at most 0.67, more preferably at most 0.50, even more preferably at most 0.25, and most preferably at most 0.15.

[0114] In some preferred embodiments of the present invention, the molar ratio between gCN and bCN in the protein solution of step a) is in the range of 0-1.0, more preferably 0.001-0.67, more preferably 0.005-0.50, even more preferably 0.01-0.25, and most preferably 0.02-0.15.

[0115] In other preferred embodiments of the invention, the molar ratio between gCN and bCN in the protein solution of step a) is in the range of 0.05-1.0, more preferably 0.1-1.0, even more preferably 0.2-1.0, and most preferably 0.3-1.0.

[0116] In another preferred embodiment of the invention, the molar ratio between gCN and bCN in the protein solution of step a) is in the range of 0.02-1.0, more preferably 0.05-0.6, and most preferably 0.08-0.4.

[0117] In some preferred embodiments of the invention, the protein solution in step a) does not contain gCN. This is typically the case, for example, if bCN has already been prepared by fermentation.

[0118] In some preferred embodiments of the present invention, the temperature of the protein solution in step a) is at most 64°C, more preferably at most 30°C, even more preferably at most 20°C, and most preferably at most 15°C.

[0119] In another preferred embodiment of the invention, the temperature of the protein solution in step a) is in the range of 0°C-64°C, more preferably 0°C-30°C, even more preferably 0°C-20°C, and most preferably 1°C-15°C.

[0120] In some preferred embodiments of the invention, providing the protein solution involves subjecting the protein feed to one or more of the following: i) Regarding the enrichment of bCN, ii) Demineralization, for example by dialysis, cation exchange and / or electrodialysis, and iii) Add a Ca-binding chelating agent.

[0121] Enrichment of bCN requires at least enrichment relative to the total protein in the protein feed, and may also involve enrichment relative to the total weight of the protein feed.

[0122] If a Ca-binding chelating agent is used, it is preferably a food-acceptable chelating agent.

[0123] The protein feed can be, for example, an aqueous liquid rich in β-casein prepared according to WO 2014 / 114709 A2, US 20070104847 A1, or WO2017220697, which, regarding suitable protein feeds, are incorporated herein by reference. These protein feeds must undergo an additional demineralization step to achieve calcium and magnesium content relative to the β-casein and γ-casein content required by the present invention.

[0124] In some preferred embodiments of the invention, the production of protein feed involves microfiltration of a liquid containing casein micelles, preferably skim milk or a dispersion of casein micelles, using an MF membrane capable of retaining casein micelles but allowing bCN permeation.

[0125] In some preferred embodiments of the invention, step b) involves heating the protein solution from step a) to a temperature of at least 70°C, more preferably at least 72°C, and most preferably at least 74°C. Preferably, the heat treatment has a duration sufficient to kill at least some of the bacteria in ps.

[0126] In another preferred embodiment of the invention, step b) involves heating the protein solution from step a) to a temperature in the range of 70°C-180°C, more preferably 72°C-160°C, and most preferably 74°C-155°C.

[0127] In even more preferred embodiments of the invention, step b) involves heating the protein solution from step a) to a temperature in the range of 70°C-90°C, more preferably 72°C-80°C, and most preferably 74°C-78°C.

[0128] Preferably, step b) involves maintaining the heated protein solution at the desired temperature for 1 second to 10 minutes, more preferably 5 seconds to 5 minutes, and most preferably a duration of 10 seconds to 1 minute.

[0129] In some preferred embodiments of the invention, the heat treatment in step b) results in a reduction in colony-forming units at a level equal to or greater than the level of heating to 70°C and holding for 10 seconds.

[0130] In other preferred embodiments of the invention, the heat treatment in step b) results in a reduction in colony-forming units at a level equal to or greater than the level of heating to 72°C and holding for 15 seconds.

[0131] In other preferred embodiments of the invention, the heat treatment in step b) results in a reduction in colony-forming units at a level equal to or greater than the level of heating to 72°C and holding for 15 seconds.

[0132] In other preferred embodiments of the invention, the heat treatment in step b) results in a reduction in colony-forming units at a level equal to or greater than the level of heating to 74°C and holding for 15 seconds.

[0133] In some preferred embodiments of the invention, step b) involves heat sterilizing the protein solution.

[0134] Following heat treatment, step b) typically includes an additional sub-step of cooling the heat-treated protein solution, preferably to a temperature of up to 20°C.

[0135] Step c) is optional, meaning that some preferred embodiments include step c) involving drying and heat treatment of the protein solution or its protein concentrate, while other preferred embodiments do not include step c).

[0136] Therefore, in some preferred embodiments of the present invention, the method of the present invention includes step c), preferably wherein the drying in step c) involves spray drying.

[0137] In the context of this invention, the term "protein concentrate" in the context of step c) refers to a liquid composition wherein at least the protein is derived from a heat-treated protein solution obtained in step b), but the liquid composition has a higher protein content relative to total solids or relative to total weight compared to the heat-treated protein solution. Preferably, substantially all the solids of the protein concentrate are derived from the heat-treated protein solution obtained in step b). Such a protein concentrate can be prepared, for example, by ultrafiltration, nanofiltration, reverse osmosis, or evaporation.

[0138] Another aspect of the invention relates to a process for producing a food containing bCN and optionally also containing gCN or its protein hydrolysates, the method comprising the following steps: I) Provides the heat-treated bCN concentrate of the present invention. II) subjecting the heat-treated bCN concentrate to further processing, such as involving the hydrolysis of bCN and / or mixing with other components and / or filling the heat-treated bCN concentrate into a suitable container.

[0139] Preferably, the food is infant formula, follow-up formula, growing-up formula, or a high-bCN beverage.

[0140] In some preferred embodiments of the invention, the food is preferably a liquid food, preferably has a pH in the range of 6-8, and more preferably in the range of 6.4-7.5.

[0141] In the context of this invention, a high-bCN beverage is a beverage containing at least 2% w / w of bCN.

[0142] The high-bCN beverage preferably contains at least 4% w / w, more preferably at least 6% w / w, and even more preferably at least 8% w / w of bCN.

[0143] High-bCN beverages preferably contain a amount of bCN in the range of 2% w / w-12% w / w, more preferably at least 4% w / w-11% w / w, and even more preferably at least 6% w / w-11% w / w.

[0144] In some preferred embodiments of the present invention, the hbc of the present invention is the sole protein source for high bCN beverages.

[0145] Generally preferred, high-bCN liquid foods have a molar ratio between at most 6, more preferably at most 4, even more preferably at most 3, and most preferably at most 2.5 of the following: The total content of calcium and magnesium, and The total content of bCN and gCN.

[0146] For example, it is preferable that the high bCN liquid food has a molar ratio between at most 2.0, more preferably at most 1.6, even more preferably at most 1.3, and most preferably at most 1.0: The total content of calcium and magnesium, and The total content of bCN and gCN.

[0147] In the context of this invention, the term "liquid food" refers to a composition containing a combination of liquid and solid or semi-solid particles such as protein particles. Thus, "liquid" can be a suspension or even a slurry. However, "liquid" is preferably pumpable.

[0148] In some preferred embodiments of the invention, the liquid food contains bCN, the amount of which is preferably in the range of 2% w / w-15% w / w, more preferably 5% w / w-14% w / w, and most preferably 8% w / w-13% w / w.

[0149] The high-bCN beverage of the present invention preferably contains one or more of the other ingredients described in WO 2020 / 002450 A1, and preferably in the amounts described in WO 2020 / 002450 A1.

[0150] In some preferred embodiments of the invention, the food is an infant formula, which preferably contains an amount of bCN in the range of 1.0% w / w to 12% w / w relative to total solids, more preferably 1.3% w / w to 8% w / w relative to total solids, and most preferably 1.5% w / w to 6% w / w relative to total solids.

[0151] The inventors have observed that the hbc of the present invention has improved bioavailability compared to existing bCN products, and this makes the hbc of the present invention suitable for, for example, infant nutrition and other nutritional applications.

[0152] In some preferred embodiments of the invention, the food, preferably infant formula, is produced by drying and blending powdered ingredients.

[0153] In the context of this invention, the term "dry blending" refers to a method of preparing powdered food, wherein the individual ingredients are provided in dry form, typically as powder, and then blended under dry conditions, i.e., without the addition of water or water-rich ingredients, to form the final food. Dry blending is typically carried out under very clean conditions, usually aseptically, and therefore does not require pasteurization heat treatment after dry blending. Furthermore, unlike wet blending methods, dry blending does not require a high-energy drying step to remove water. Wet-mixing also requires a second pasteurization step, which tends to cause further protein denaturation and degradation of other heat-sensitive components of the powdered food. The higher content of denatured proteins in powdered foods prepared by wet blending can also cause increased viscosity when the powdered food is resuspended in water, and may make the resulting liquid beverage less drinkable.

[0154] In the context of this invention, the term "infant formula" refers to a nutritionally complete food for infants aged 0-12 months, preferably conforming to Part 107 (Infant Formula) D (Nutritional Requirements) of Title 21, Chapter 1, Subsection B of the United States Federal Regulations; Section 107.100 Nutritional Specifications, which came into effect on 1 April 2015; or to European Union Regulation 609 / 2013, which specifies standards for foods for special medical purposes.

[0155] In some preferred embodiments of the invention, the heat-treated bCN concentrate is used in an amount sufficient to provide at least 50% w / w, more preferably at least 70% w / w, even more preferably at least 80% w / w, and most preferably at least 90% w / w of the bCN in the food.

[0156] Another aspect of the present invention relates to a food product obtainable by the process of the present invention.

[0157] Another aspect of the invention relates to the use of the heat-treated bCN concentrate of the invention as a food ingredient, preferably for the production of one or more of the foods mentioned herein, preferably using one or more features of the process mentioned above.

[0158] Further preferred embodiments of the invention are described in the following numbered embodiments.

[0159] Scheme 1. A heat-treated β-casein concentrate (HBC) comprising: The total content of β-casein (bCN) and γ-casein (gCN) relative to the weight of hbc is at least 9% w / w. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein. The molar ratio between the following items is at most 6.0: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

[0160] Scheme 2. The heat-treated bCN concentrate according to Scheme 1, in powder form, preferably has a total solids content of at least 90% w / w, more preferably at least 92% w / w, even more preferably at least 94% w / w, and most preferably at least 95% w / w.

[0161] Scheme 3. The heat-treated bCN concentrate according to Scheme 1, in liquid form, preferably has a total solids content in the range of 9% w / w-50% w / w, more preferably 9% w / w-40% w / w, even more preferably 10% w / w-30% w / w, and most preferably 11% w / w-25% w / w.

[0162] Scheme 4. The heat-treated bCN concentrate according to any one of the aforementioned schemes has a total bCN and gCN content of at least 9% w / w relative to the weight of hbc, more preferably at least 9.5% w / w, even more preferably at least 10% w / w, and most preferably at least 11% w / w relative to the weight of hbc.

[0163] Scheme 5. The heat-treated bCN concentrate according to any one of the aforementioned schemes has a total bCN and gCN content of 9% w / w-25% w / w, more preferably 9% w / w-20% w / w, even more preferably 9.5% w / w-16% w / w, and most preferably 10% w / w-14% w / w relative to the weight of hbc.

[0164] Scheme 6. The heat-treated bCN concentrate according to any one of the aforementioned schemes has a total bCN and gCN content of 9% w / w-14% w / w, more preferably 9% w / w-13% w / w, even more preferably 9.5% w / w-12% w / w, and most preferably 9.5% w / w-11% w / w relative to the weight of hbc.

[0165] Scheme 7. The heat-treated bCN concentrate according to any one of the aforementioned schemes has a total content of bCN and gCN of at least 40% w / w relative to the weight of hbc, more preferably at least 50% w / w, even more preferably at least 60% w / w, and most preferably at least 70% w / w relative to the weight of hbc.

[0166] Scheme 8. The heat-treated bCN concentrate according to any one of the aforementioned schemes, having a total content of bCN and gCN of at least 80% w / w relative to the weight of hbc, more preferably at least 90% w / w, even more preferably at least 95% w / w, and most preferably at least 98% w / w relative to the weight of hbc.

[0167] Scheme 9. The heat-treated bCN concentrate according to any one of the aforementioned schemes has a total bCN and gCN content of 40% w / w-98% w / w, more preferably 50% w / w-95% w / w, even more preferably 55% w / w-90% w / w, and most preferably 60% w / w-85% w / w relative to the weight of hbc.

[0168] Scheme 10. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the hbc has a molar ratio of at most 1.0 between gCN and bCN.

[0169] Scheme 11. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the molar ratio of gCN to bCN in hbc is at most 0.67, more preferably at most 0.50, even more preferably at most 0.25, and most preferably at most 0.15.

[0170] Scheme 12. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the molar ratio of gCN to bCN of hbc is 0-1.0, more preferably 0.001-0.67, more preferably 0.005-0.50, even more preferably 0.01-0.25, and most preferably 0.02-0.15.

[0171] Scheme 13. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the molar ratio between gCN and bCN of hbc is 0.05-1.0, more preferably 0.1-1.0, even more preferably 0.2-1.0, and most preferably 0.3-1.0.

[0172] Scheme 14. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the molar ratio of gCN to bCN of hbc is 0.02-1.0, more preferably 0.05-0.6, and most preferably 0.08-0.4.

[0173] Scheme 15. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the hbc does not contain gCN.

[0174] Scheme 16. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the hbc contains a total protein content of at least 50% w / w, more preferably at least 70% w / w, even more preferably at least 80% w / w, and most preferably at least 90% w / w relative to the total solids.

[0175] Scheme 17. The heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the total protein content of the hbc is in the range of 6% w / w-50% w / w, more preferably 8% w / w-40% w / w, even more preferably 10% w / w-30% w / w, and most preferably 12% w / w-20% w / w.

[0176] Scheme 18. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the total content of bCN and gCN of hbc is at least 25% w / w relative to the total protein, and more preferably at least 40% w / w relative to the total protein.

[0177] Scheme 19. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the total content of bCN and gCN of hbc is at least 50% w / w, more preferably at least 60% w / w, even more preferably at least 70% w / w, and most preferably at least 80% w / w relative to the total protein.

[0178] Embodiment 20. A heat-treated bCN concentrate according to any one of the preceding embodiments, wherein the total content of bCN and gCN of the hbc is at least 85% w / w relative to the total amount of protein, more preferably at least 90% w / w, and most preferably at least 95% w / w relative to the total amount of protein.

[0179] Scheme 21. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the total content of bCN and gCN of hbc is 40% w / w-98% w / w, more preferably 50% w / w-96% w / w, even more preferably 60% w / w-94% w / w, and most preferably 70% w / w-92% w / w relative to the total protein.

[0180] Scheme 22. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the total content of bCN and gCN of hbc is 40% w / w-80% w / w, more preferably 40% w / w-75% w / w, even more preferably 40% w / w-70% w / w, and most preferably 40% w / w-65% w / w relative to the total protein.

[0181] Scheme 23. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the total content of bCN and gCN of hbc is at least 70% w / w, more preferably at least 80% w / w, even more preferably at least 90% w / w, even more preferably at least 95% w / w, and most preferably at least 98% w / w relative to the total amount of casein.

[0182] Scheme 24. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the total content of bCN and gCN of hbc is 65% w / w-98% w / w, more preferably 70% w / w-96% w / w, even more preferably 75% w / w-94% w / w, and most preferably 80% w / w-94% w / w relative to the total amount of casein.

[0183] Scheme 25. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the hbc contains whey protein in an amount of 10% w / w to 40% w / w relative to non-bCN protein, more preferably at least 15% w / w to 35% w / w, and most preferably 20% w / w to 30% w / w relative to non-bCN protein.

[0184] Scheme 26. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the hbc contains whey protein in an amount of at least 50% w / w relative to non-bCN protein, more preferably at least 70% w / w, and most preferably at least 90% w / w relative to non-bCN protein.

[0185] Scheme 27. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the hbc contains bCN peptides in an amount of 60% w / w-90% w / w, more preferably 65% ​​w / w-85% w / w, and most preferably 70% w / w-80% w / w relative to non-bCN proteins.

[0186] Embodiment 28. A heat-treated bCN concentrate according to any one of the preceding embodiments, wherein the hbc has a molar ratio between at most 5, more preferably at most 4, even more preferably at most 3, and most preferably at most 2.5 of the following: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

[0187] Embodiment 29. A heat-treated bCN concentrate according to any one of the preceding embodiments, wherein the hbc has a molar ratio between at most 2.0, more preferably at most 1.6, even more preferably at most 1.3, and most preferably at most 1.0: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

[0188] Embodiment 30. A heat-treated bCN concentrate according to any one of the preceding embodiments, wherein the hbc has a molar ratio between at most 6, more preferably at most 5, even more preferably at most 4, even more preferably at most 3, and most preferably at most 2.5 of the following: The total content of calcium and magnesium, and The total content of bCN and gCN.

[0189] Embodiment 31. A heat-treated bCN concentrate according to any one of the preceding embodiments, wherein the hbc has a molar ratio between at most 2.0, more preferably at most 1.6, even more preferably at most 1.3, and most preferably at most 1.0: The total content of calcium and magnesium, and The total content of bCN and gCN.

[0190] Scheme 32. The heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the amount of carbohydrate contained in the hbc is at most 15% w / w, more preferably at most 10% w / w, even more preferably at most 5% w / w, and most preferably at most 1% w / w.

[0191] Scheme 33. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the hbc contains carbohydrates in an amount of up to 50% w / w relative to total solids, more preferably up to 40% w / w relative to total solids, even more preferably up to 30% w / w relative to total solids, and most preferably up to 20% w / w relative to total solids.

[0192] Scheme 34. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the hbc contains carbohydrates in an amount of up to 15% w / w relative to total solids, more preferably up to 10% w / w relative to total solids, even more preferably up to 5% w / w relative to total solids, and most preferably up to 1% w / w relative to total solids.

[0193] Scheme 35. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the amount of lactose contained in the hbc is at most 15% w / w, more preferably at most 10% w / w, even more preferably at most 5% w / w, and most preferably at most 1% w / w.

[0194] Scheme 36. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the hbc contains lactose in an amount of up to 50% w / w relative to total solids, more preferably up to 40% w / w relative to total solids, even more preferably up to 30% w / w relative to total solids, and most preferably up to 20% w / w relative to total solids.

[0195] Scheme 37. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the hbc contains lactose in an amount of up to 15% w / w relative to total solids, more preferably up to 10% w / w relative to total solids, even more preferably up to 5% w / w relative to total solids, and most preferably up to 1% w / w relative to total solids.

[0196] Embodiment 38. A heat-treated bCN concentrate according to any one of the preceding embodiments, wherein the amount of lipids contained in the hbc is at most 5% w / w, more preferably at most 2% w / w, even more preferably at most 1% w / w, and most preferably at most 0.2% w / w.

[0197] Embodiment 39. A heat-treated bCN concentrate according to any one of the preceding embodiments, wherein the hbc contains lipids in an amount of up to 15% w / w relative to total solids, more preferably up to 12% w / w relative to total solids, even more preferably up to 10% w / w relative to total solids, and most preferably up to 8% w / w relative to total solids.

[0198] Embodiment 40. A heat-treated bCN concentrate according to any one of the preceding embodiments, wherein the hbc contains lipids in an amount of up to 5% w / w relative to total solids, more preferably up to 2% w / w relative to total solids, even more preferably up to 1% w / w relative to total solids, and most preferably up to 0.2% w / w relative to total solids.

[0199] Scheme 41. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the pH of the hbc is in the range of 6-9, more preferably 6.1-8.0, even more preferably 6.3-7.5 and most preferably 6.5-7.2.

[0200] Scheme 42. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the pH of the hbc is in the range of 6.5-9, more preferably 6.7-8.0, even more preferably 7.0-8.0 and most preferably 7.1-8.0.

[0201] Scheme 43. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the hbc has: pH in the range of 6-7.0, and The molar ratios between at most 5, more preferably at most 4, even more preferably at most 3, and most preferably at most 2.5 of the following: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

[0202] Scheme 44. A heat-treated bCN concentrate according to any one of the aforementioned schemes, wherein the hbc has: pH in the range of 6-7.0, and The molar ratio between the following items is at most 2.0, more preferably at most 1.6, even more preferably at most 1.3, and most preferably at most 1.0: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

[0203] Scheme 45. A heat-treated bCN concentrate according to any one of the preceding schemes, wherein the hbc has: pH in the range of 6-7.0, and The molar ratios between at most 5, more preferably at most 4, even more preferably at most 3, and most preferably at most 2.5 of the following: The total content of calcium and magnesium, and The total content of bCN and gCN.

[0204] Scheme 46. A heat-treated bCN concentrate according to any one of the preceding schemes, wherein the hbc has: pH in the range of 6-7.0, and The molar ratio between the following items is at most 2.0, more preferably at most 1.6, even more preferably at most 1.3, and most preferably at most 1.0: The total content of calcium and magnesium, and The total content of bCN and gCN.

[0205] Scheme 47. The heat-treated bCN concentrate according to any one of the aforementioned schemes has a colony-forming unit content of up to 10,000 CFU / g total solids, more preferably up to 5,000 CFU / g total solids, even more preferably up to 1,000 CFU / g total solids, and most preferably up to 100 CFU / g total solids.

[0206] Scheme 48. The heat-treated bCN concentrate according to any one of the aforementioned schemes is sterile and preferably heat-sterilized.

[0207] Embodiment 49. A heat-treated bCN concentrate according to any one of the preceding embodiments, which is in powder form, preferably prepared by spray drying, and has a concentration of at least 0.20 g / cm³. 3 More preferably at least 0.22 g / cm³ 3 And most preferably at least 0.24 g / cm³ 3 The packing density within the range.

[0208] Embodiment 50. A heat-treated bCN concentrate according to any one of the preceding embodiments, which is in powder form, preferably prepared by spray drying, and having a concentration of 0.20 g / cm³. 3 -1.0 g / cm 3 More preferably 0.21 g / cm 3 -0.6 g / cm 3 And the most preferred value is 0.22 g / cm³. 3 -0.4 g / cm 3 The packing density within the range.

[0209] Scheme 51. A heat-treated bCN concentrate according to any one of Schemes 1-50, having a viscosity of [viscosity] at 25°C and 145 s when adjusted to a protein content of 10.0% w / w. -1 The shear rate is at most 30 mPa*s, more preferably at 145 s at 25°C. -1 The shear rate is at most 25 mPa*s, and even more preferably at 25°C for 145 s. -1 The shear rate is at most 22 mPa*s, and most preferably at 25°C for 145 s. -1 The shear rate is at most 20 mPa*s.

[0210] Embodiment 52. A method for producing a heat-treated β-casein concentrate according to any one of embodiments 1-51, said method comprising the steps of: a) Provide a protein solution (ps), said protein solution (ps) having: The total content of β-casein (bCN) and γ-casein (gCN) relative to the weight of the protein solution is at least 9% w / w. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein. Preferably, the molar ratio between at most 6.0 of the following items is: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN, and pH range of 6-9 b) Heat the protein solution from step a) to a temperature of at least 65°C, and c) Optionally, the heat-treated protein solution or its protein concentrate is dried.

[0211] Scheme 53. The method according to Scheme 52, wherein the total content of bCN and gCN of the protein solution (ps) in step a) is at least 9% w / w relative to the weight of ps, more preferably at least 9.5% w / w, even more preferably at least 10% w / w, and most preferably at least 11% w / w relative to the weight of ps.

[0212] Scheme 54. The method according to Scheme 52 or 53, wherein the total content of bCN and gCN of the protein solution in step a) is 9% w / w-25% w / w, more preferably 9% w / w-20% w / w, even more preferably 9.5% w / w-16% w / w, and most preferably 10% w / w-14% w / w relative to the weight of ps.

[0213] Scheme 55. The method according to any one of Schemes 52-54, wherein the total content of bCN and gCN of the protein solution in step a) is 9% w / w-14% w / w, more preferably 9% w / w-13% w / w, even more preferably 9.5% w / w-12% w / w, and most preferably 9.5% w / w-11% w / w relative to the weight of ps.

[0214] Scheme 56. The method according to any one of Schemes 52-55, wherein the protein solution in step a) has a molar ratio of at most 1.0 between gCN and bCN.

[0215] Scheme 57. The method according to any one of Schemes 52-56, wherein the molar ratio between gCN and bCN of the protein solution in step a) is at most 0.67, more preferably at most 0.50, even more preferably at most 0.25, and most preferably at most 0.15.

[0216] Scheme 58. The method according to any one of Schemes 52-57, wherein the molar ratio between gCN and bCN of the protein solution in step a) is 0-1.0, more preferably 0.001-0.67, more preferably 0.005-0.50, even more preferably 0.01-0.25, and most preferably 0.02-0.15.

[0217] Scheme 59. The method according to any one of Schemes 52-58, wherein the molar ratio between gCN and bCN in the protein solution of step a) is 0.05-1.0, more preferably 0.1-1.0, even more preferably 0.2-1.0, and most preferably 0.3-1.0.

[0218] Scheme 60. The method according to any one of Schemes 52-59, wherein the molar ratio between gCN and bCN in the protein solution of step a) is 0.02-1.0, more preferably 0.05-0.6, and most preferably 0.08-0.4.

[0219] Scheme 61. The method according to any one of Schemes 52-60, wherein the protein solution in step a) does not contain gCN.

[0220] Scheme 62. The method according to any one of Schemes 52-61, wherein the protein solution in step a) has a temperature of up to 64°C, more preferably up to 30°C, even more preferably up to 20°C, and most preferably up to 15°C.

[0221] Scheme 63. The method according to any one of Schemes 52-62, wherein the protein solution in step a) has a temperature of 0°C-64°C, more preferably 0°C-30°C, even more preferably 0°C-20°C, and most preferably 1°C-15°C.

[0222] Scheme 64. The method according to any one of Schemes 52-63, wherein providing the protein solution involves subjecting the protein feed to one or more of the following: i) Regarding the enrichment of bCN, ii) Demineralization, for example by dialysis, cation exchange and / or electrodialysis, and iii) Add a Ca-binding chelating agent.

[0223] Embodiment 65. The method according to Embodiment 64, wherein the production of the protein feed involves microfiltration (MF) of a liquid containing casein micelles using an MF membrane capable of retaining casein micelles but allowing bCN permeation, preferably wherein the liquid is skim milk or a dispersion of casein micelles.

[0224] Scheme 66. The method according to any one of Schemes 52-65, wherein step b) involves heating the protein solution of step a) to a temperature of at least 70°C, more preferably at least 72°C, and most preferably at least 74°C.

[0225] Scheme 67. The method according to any one of Schemes 52-66, wherein step b) involves heating the protein solution of step a) to a temperature in the range of 70°C-180°C, more preferably 72°C-160°C, and most preferably 74°C-155°C.

[0226] Scheme 68. The method according to any one of Schemes 52-67, wherein step b) involves heating the protein solution of step a) to a temperature in the range of 70°C-90°C, more preferably 72°C-80°C, and most preferably 74°C-78°C.

[0227] Scheme 69. The method according to any one of Schemes 52-68, wherein step b) involves maintaining the heated protein solution at the desired temperature for a duration of 1 second to 10 minutes, more preferably 5 seconds to 5 minutes, and most preferably 10 seconds to 1 minute.

[0228] Scheme 70. The method according to any one of Schemes 52-69, wherein the heat treatment in step b) results in a reduction in colony-forming units at a level equal to or greater than the level of heating to 70°C and holding for 10 seconds.

[0229] Scheme 71. The method according to any one of Schemes 52-70, wherein the heat treatment in step b) results in a reduction in colony-forming units at a level equal to or greater than the level of heating to 72°C and holding for 15 seconds.

[0230] Scheme 72. The method according to any one of Schemes 52-71, wherein the heat treatment in step b) results in a reduction in colony-forming units at a level equal to or greater than the level of heating to 74°C and holding for 15 seconds.

[0231] Scheme 73. The method according to any one of Schemes 52-72, wherein step b) involves heat sterilizing the protein solution.

[0232] Scheme 74. The method according to any one of Schemes 52-73, comprising step c), and preferably wherein the drying in step c) involves spray drying.

[0233] Scheme 75. A process for producing a food comprising bCN and optionally also comprising gCN or its protein hydrolysates, the method comprising the following steps: I) Provide a heat-treated bCN concentrate according to one or more of the embodiments 1-51, II) subject the heat-treated bCN concentrate to further processing, such as involving the hydrolysis of bCN and / or mixing with other components and / or filling the heat-treated bCN concentrate into a suitable container.

[0234] Scheme 76. The process according to Scheme 75, wherein the food is an infant formula, the infant formula preferably containing an amount of bCN in the range of 1.0% w / w to 12% w / w relative to total solids, more preferably 1.3% w / w to 8% w / w relative to total solids, and most preferably in the range of 1.5% w / w to 6% w / w relative to total solids.

[0235] Scheme 77. The process according to Scheme 75 or 76, wherein the food, preferably infant formula, is produced by drying and blending powder components.

[0236] Scheme 78. The process according to Scheme 75, wherein the food is a liquid food, and the liquid food preferably has a pH in the range of 6-8.

[0237] Scheme 79. The process according to Scheme 78, wherein the liquid food contains an amount of bCN in the range of 2% w / w-15% w / w, more preferably 5% w / w-14% w / w, and most preferably 8% w / w-13% w / w.

[0238] Scheme 80. The process according to any one of Schemes 75-79, wherein the heat-treated bCN concentrate is used in an amount sufficient to provide at least 50% w / w, more preferably at least 70% w / w, even more preferably at least 80% w / w, and most preferably at least 90% w / w of the bCN of the food.

[0239] Implementation scheme 81. A food product obtainable by the process described in one or more of implementation schemes 75-80.

[0240] Implementation scheme 82. Use of the heat-treated bCN concentrate according to one or more of implementation schemes 1-51 as a food ingredient. Example

[0241] Analytical methods The parameters mentioned in this application were determined using the analysis mentioned in WO 2020 / 002450 A1 or described below.

[0242] Analysis A: Determination of the relative concentrations of γ-casein and β-casein individually: LCMS The relative concentrations of individual γ-casein and β-casein were determined by liquid chromatography-high resolution mass spectrometry (LC-HRMS) using the method described by Nilsson et al. (2020) Journal of Dairy Science, 103 (2020), pp. 6858-6868 and quantified using area under the curve.

[0243] The powdered sample was dissolved in trisodium citrate (100 mM; Merck, Darmstadt, Germany) at a concentration of 20 g / L and treated at 30 °C for 60 min with 20 μL of dithioerythritol (0.5 M; Sigma Aldrich, St. Louis, MO, USA) and 1 mL of urea (6 M; Sigma Aldrich) and trisodium citrate (100 mM; Merck, Darmstadt, Germany) buffer. The sample was centrifuged for 10 min (at 5 °C, 16100 xg), and then 5 μL of the supernatant was injected into a 1290 LC (Agilent Technologies, Santa Clara, CA) linked to a 6230 HRMS, using a Poroshell 120 SB-C18 2.1 × 150 mm, 2.7 mm (Agilent Technologies) column at 40 °C with UV detection at 214 nm. To separate CN and whey protein, two mobile phases were used: A contained 0.05% (v / v) trifluoroacetic acid (TFA; Thermo Fisher Scientific) in MilliQ water, and B contained 0.1% (v / v) TFA in acetonitrile (Rathburn, Walkerburn, UK). The gradient had the following composition: 15% B at 1 min, 34% B at 4 min, and 47% B at 19 min.

[0244] Data from UV chromatograms and total ion chromatograms were analyzed using MassHunter and Profinder software (Agilent Technologies) to obtain the relative concentration of individual γ-casein in each total protein.

[0245] The concentrations of individual γ-casein and β-casein can be converted to molar content using the molecular weight of the protein. If only the total weight-based content of γ-casein is provided, the molar content of gCN is determined based on the average molecular weight of the gCN substance (14600 g / mol). The molecular weights of bovine γ-1-casein, γ-2-casein, and γ-3-casein are approximately 20.4 kDa / mol, 11.8 kDa / mol, and 11.6 kDa / mol, respectively.

[0246] Example 1: The bCN problem and the solution of the present invention The inventors have experienced several instances where protein solutions containing high concentrations of β-casein were difficult to process and handle, and have observed sudden clogging of processing equipment or undesirable sensory changes in the final product. These problems occur both during the production of specialized bCN ingredient powders, for example, for use in infant formula or high-protein beverage products, and in subsequent use of the bCN ingredient powders in such applications.

[0247] The inventors have studied these problems and have surprisingly discovered that these problems can be reduced or even avoided by controlling the molar ratio between the combined content of calcium and magnesium and the combined content of β-casein and γ-casein.

[0248] The following experiments A and B illustrate both the problem and the solution.

[0249] Experiment A: β-casein concentrate is produced according to the method given below: β-casein concentrate was produced using bovine skim milk as feedstock by combining warm and cold microfiltration in a given sequence. A given volume of skim milk was subjected to continuous microfiltration at 50°C using a spiral-wound membrane of type FR6338 (nominal cutoff 800,000 Daltons) from Synder Filtration, Vacaville, California, US, with a concentration factor of 2.5. Permeate from the microfiltration process was continuously directed to a parallel ultrafiltration process (HFK-328 6338, from KochMembrane Systems, Wilmington, Massachusetts, US, nominal cutoff 5,000 Daltons), permeate from the ultrafiltration process was continuously directed to a reverse osmosis filtration process, and permeate from the reverse osmosis process was directed back to the microfiltration residue for percolation. 200% percolation was performed, meaning the volume of reverse osmosis permeate used for percolation was twice the volume of skim milk supplied to the microfiltration process, and the process was operated at 50°C. The processed microfiltration residue was subjected to continuous heat treatment at 74°C for 15 seconds, cooled to 6°C, and collected in a tank for storage at 6°C for 24 hours.

[0250] Using a spiral-wound FR6338 type membrane with a nominal cutoff of 800,000 Daltons from Synder Filtration, Vacaville, California, US, a given volume of stored microfiltration residue was subjected to continuous microfiltration at 6°C with a concentration factor of 1.1. The permeate from the microfiltration process was then directed to a parallel ultrafiltration process (HFK-328 6338, from Koch Membrane Systems, Wilmington, Massachusetts, US, with a nominal cutoff of 5,000 Daltons), involving 35% percolation using reverse osmosis permeate, concentrating the microfiltration permeate to approximately 12% dry matter in the ultrafiltration residue, and continuously returning the permeate from the ultrafiltration process to the microfiltration residue for further percolation. A 500% percolation was performed, meaning the volume of ultrafiltration permeate used for percolation was five times the volume of the microfiltration residue solution supplied to the microfiltration process. When the filtration process is complete, approximately 1000 liters of ultrafiltration residue are collected, with a dry matter content of 10.6% w / w and a protein content of 10.1% w / w.

[0251] The ultrafiltration residue was subjected to a separate ultrafiltration process (HFK-328 6338, from Koch Membrane Systems, Wilmington, Massachusetts, USA, nominal cutoff 5,000 Daltons) for concentration, yielding approximately 500 kg of liquid β-casein concentrate with a dry matter content of 15.2% w / w, a protein content of 14.8% w / w, and a total protein content of 73.1% w / w for both β-casein and γ-casein (absolute β-casein + γ-casein content of 10.8% w / w). The general composition of the β-casein concentrate is given in Table 1.1. Table 1.1: General composition of β-casein concentrate from Experiment A

[0252] Heat treatment of β-casein concentrate: A solution of β-casein concentrate was heat-treated to 72°C using a standard plate heat exchanger and held at that temperature for 15 seconds. However, shortly after the start of the heat treatment process, the pressure in the heat exchanger suddenly increased to above 8 bar, halting the process; that is, heat treatment under the given conditions was impossible.

[0253] Experiment B: Based on the principles of Experiment A, a similar β-casein concentrate was produced to provide a liquid β-casein concentrate with a dry matter content of 5.1% and a protein content of 5.0%.

[0254] The liquid β-casein concentrate was then subjected to cation exchange chromatography using 360 liters of IMAC HP333 resin (DuPont, Edina, Minnesota, US) contained in a column, with the aim of removing calcium.

[0255] Prior to the cation exchange operation, the resin in the column is fully regenerated, leaving resin in sodium form. The flow product effluent from the column loaded with ultrafiltration residue is collected, yielding approximately 1000 liters of cation-exchanged liquid β-casein concentrate, some of which is diverted for discharge. For the cation exchange operation, virtually no protein loss was observed; that is, the relative composition of the protein components, including the content of β-casein relative to total protein, remained the same before and after the cation exchange operation. The general composition of the liquid β-casein concentrate before and after cation exchange is given in Table 1.2 below.

[0256] Table 1.2

[0257] The cation-exchanged liquid β-casein concentrate was further concentrated by ultrafiltration using a membrane with a nominal cutoff of 5,000 Daltons, yielding approximately 210 kg of liquid β-casein concentrate with a dry matter content of 16.9%, a protein content of 16.2%, and a total protein content of 72.3% (absolute β-casein + γ-casein content of 11.7%). The composition of the concentrated cation-exchanged β-casein concentrate is shown in Table 1.3.

[0258] Table 1.3 General composition of concentrated decalcified β-casein concentrate

[0259] Heat treatment of β-casein concentrate: The solution of β-casein concentrate was heated using a standard plate heat exchanger in a manner similar to that in Experiment A. The heat treatment process was completed without any problematic pressure increases during operation.

[0260] The molar ratio between the combined content of calcium and magnesium and the combined content of β-casein and γ-casein was approximately 7.24 in the product of Experiment A and less than 0.5 in Experiment B. Other experiments conducted by the inventors have shown that increasing the content of monovalent metal salts without decreasing the combined content of calcium and magnesium does not solve the problem identified by the inventors.

[0261] The proteins in the products of experiments A and B were mainly composed of β-casein, γ-casein, and whey protein. β-casein and γ-casein contributed more than 95% of the total casein.

[0262] in conclusion: The inventors have discovered that the brief heat treatment that occurs during the pasteurization of liquid β-casein concentrates with a β-casein and γ-casein content of 9% w / w or higher is challenging and can lead to stress problems and even system failure. However, the inventors have found that this problem can be surprisingly solved by controlling the molar ratio between the combined content of calcium and magnesium and the combined content of β-casein and γ-casein prior to pasteurization.

[0263] If the process flow used contains a high concentration of β-casein, similar problems as experienced in Experiment A may occur during the production of high-protein beverages based on bCN as the main protein source or during the production of infant formula. The inventors have observed indications that these problems can also be remedied by controlling the molar ratio between the combined content of calcium and magnesium and the combined content of β-casein and γ-casein in the bCN ingredients used.

[0264] Furthermore, the present invention enables efficient heat treatment during the production of the bCN components of the invention, thereby reducing microorganisms without compromising the functionality of the components. Therefore, the present invention opens up a method for the production of heat-treated bCN concentrates, which are, for example, in liquid or powder form, have very low microbial content, for example less than 10,000 CFU / g total solids, more preferably less than 1,000 CFU / g total solids, and most preferably the heat-treated bCN concentrates are sterile.

[0265] Example 2 - High-protein beverage based on β-casein This embodiment discloses the preparation of 24 samples of high-protein model beverages (approximately 5.5%, 11%, and 16% protein) based on β-casein as the primary protein source, and the benefits of using the novel β-casein concentrate of this invention as a β-casein source. The beverage samples had a pH of 6.5 or 7.0 and had undergone UHT treatment or pasteurization.

[0266] Materials and Methods (The present invention) bCN-low Ca powder was prepared by spray drying the final product of Experiment B in Example 1. bCN-normal Ca powder was prepared by spray drying the unheated product of Experiment A in Example 1 (reference). NaOH and glucono delta-lactone (GDL) were purchased from Sigma Aldrich, Denmark. Demineralized water The beverage samples according to Tables 2.1, 2.2, 2.3, and 2.4 were prepared by mixing an appropriate amount of protein powder with approximately 90% of the required demineralized water, adjusting the pH to the target pH using a dilution solution of NaOH or GDL, and finally adding the remaining demineralized water to achieve the target weight for the beverage batch. The resulting mixture was allowed to hydrate at 6°C for 2 hours.

[0267] Table 2.1: Composition of bCN beverage sample at pH 6.5. Paste. = Pasteurized at 72°C for 15 seconds.

[0268]

[0269] *) Until the final sample weight is 1000 g Table 2.2: Composition of bCN beverage samples at pH 7.0. Paste. = Pasteurized at 72°C for 15 seconds.

[0270]

[0271] *) Until the final sample weight is 1000 g Table 2.3: Composition of bCN beverage sample at pH 6.5. UHT = UHT-type heat treatment at 143°C for 4 seconds.

[0272]

[0273] *) Until the final sample weight is 1000 g Table 2.4: Composition of bCN beverage sample at pH 7.0. UHT = UHT-type heat treatment at 143°C for 4 seconds.

[0274]

[0275] *) Until the final sample weight is 1000 g The beverage samples were heat-treated under the following conditions: a) 143°C for 4 seconds (UHT), comprising preheating the unheated sample to 50°C, heating the preheated sample to 143°C via direct steam injection, holding the sample at that temperature for 4 seconds, rapidly cooling the sample to 50°C, and subsequently cooling the heat-treated sample to 10°C, or b) 72°C, held for 15 seconds (pasteurization), using a plate heat exchanger (manufacturer: OMVE HTST / UHT pilot plant HT320-20) equipped with a 10-micron adhesive microfiber filter element, code 12-57-60k (title filter); and then the heated sample was cooled to 10°C.

[0276] The heat-treated beverage sample was aseptically filled into a 100 mL sterile bottle.

[0277] Characterization: Beverage samples treated with UHT were stored at ambient temperature for 2 hours, 7 days, 14 days, 1 month, and 6 months, and then characterized by visual observation and viscosity measurement (using an Anton Paar rheometer according to Example 1.8 of WO2020002450A1).

[0278] Pasteurized beverage samples were stored at 5°C for 2 hours, 7 days, and 14 days, and then characterized by visual observation and viscosity measurement (using an Anton Paar rheometer according to Example 1.8 of WO2020002450A1).

[0279] Results / Conclusions The results of Example 1 and other findings of the inventors indicate that, compared to beverages produced using conventional β-casein ingredients, the β-casein concentrate-based high-protein beverage of the present invention is less prone to system failures during production and will have a smaller tendency for undesirable thickening during storage.

[0280] Undesirable thickening of beverage products, such as at room temperature, is often seen by consumers as a product defect, and therefore reducing or even avoiding thickening problems is beneficial.

Claims

1. A heat-treated β-casein concentrate (HBC), comprising: The total content of β-casein (bCN) and γ-casein (gCN) relative to the weight of the hbc is at least 9% w / w. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein. The molar ratio between the following items is at most 6.0: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN Preferably, the pH is in the range of 6-9, more preferably 6.1-8.0, even more preferably 6.3-7.5, and most preferably 6.5-7.

2. Preferably, the molar ratio between gCN and bCN is at most 1.0, and Preferably, the content is at most 10,000 CFU / g total solids, more preferably at most 5,000 CFU / g total solids, even more preferably at most 1,000 CFU / g total solids, and most preferably at most 100 CFU / g total solids.

2. The heat-treated bCN concentrate according to claim 1, in powder form, preferably having a total solids content of at least 90% w / w, more preferably at least 92% w / w, even more preferably at least 94% w / w, and most preferably at least 95% w / w.

3. The heat-treated bCN concentrate according to claim 1, in liquid form, preferably having a total solids content in the range of 9% w / w-50% w / w, more preferably 9% w / w-40% w / w, even more preferably 10% w / w-30% w / w, and most preferably 11% w / w-25% w / w.

4. The heat-treated bCN concentrate according to any one of the preceding claims, having a total bCN and gCN content of 9% w / w-25% w / w, more preferably 9% w / w-20% w / w, even more preferably 9.5% w / w-16% w / w, and most preferably 10% w / w-14% w / w relative to the weight of the hbc.

5. The heat-treated bCN concentrate according to any one of the preceding claims, having a total bCN and gCN content of 40% w / w-98% w / w, more preferably 50% w / w-95% w / w, even more preferably 55% w / w-90% w / w, and most preferably 60% w / w-85% w / w relative to the weight of the hbc.

6. The heat-treated bCN concentrate according to any one of the preceding claims, wherein the molar ratio of gCN to bCN of hbc is at most 0.67, more preferably at most 0.50, even more preferably at most 0.25, and most preferably at most 0.

15.

7. The heat-treated bCN concentrate according to any one of the preceding claims, wherein the molar ratio of gCN to bCN in hbc is 0.02-1.0, more preferably 0.05-0.6, and most preferably 0.08-0.

4.

8. The heat-treated bCN concentrate according to any one of the preceding claims, wherein the total content of bCN and gCN of the hbc is at least 50% w / w relative to the total protein, more preferably at least 60% w / w, even more preferably at least 70% w / w, and most preferably at least 80% w / w relative to the total protein.

9. The heat-treated bCN concentrate according to any one of the preceding claims, wherein the hbc contains whey protein in an amount of at least 50% w / w relative to non-bCN protein, more preferably at least 70% w / w, and most preferably at least 90% w / w relative to non-bCN protein.

10. The heat-treated bCN concentrate according to any one of the preceding claims, wherein the hbc has a molar ratio between at most 5, more preferably at most 4, even more preferably at most 3, and most preferably at most 2.5 of the following: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN.

11. The heat-treated bCN concentrate according to any one of the preceding claims, wherein the hbc has a molar ratio between at most 6, more preferably at most 5, even more preferably at most 4, even more preferably at most 3, and most preferably at most 2.5 of the following: The total content of calcium and magnesium, and The total content of bCN and gCN.

12. The heat-treated bCN concentrate according to any one of the preceding claims, in powder form, preferably prepared by spray drying, and having a concentration of at least 0.20 g / cm³. 3 More preferably at least 0.22 g / cm³ 3 And most preferably at least 0.24 g / cm³ 3 The packing density within the range.

13. The heat-treated bCN concentrate according to any one of the preceding claims, having a viscosity of [viscosity value] at 25°C and 145 s when adjusted to a protein content of 10.0% w / w. -1 The shear rate is at most 30 mPa*s, more preferably at 145 s at 25°C. -1 The shear rate is at most 25 mPa*s, and even more preferably at 25°C for 145 s. -1 The shear rate is at most 22 mPa*s, and most preferably at 25°C for 145 s. -1 The shear rate is at most 20 mPa*s.

14. A method for producing a heat-treated β-casein concentrate according to any one of claims 1-13, the method comprising the following steps: a) Provide a protein solution, said protein solution having: The total content of β-casein and γ-casein is at least 9% w / w relative to the weight of the protein solution. The total content of bCN and gCN is at least 65% w / w relative to the total amount of casein. Preferably, the molar ratio between at most 6.0 of the following items is: The total content of casein-bound calcium and casein-bound magnesium, and The total content of bCN and gCN, and pH range of 6-9 b) Heat the protein solution from step a) to a temperature of at least 65°C, and c) Optionally, the heat-treated protein solution or its protein concentrate is dried.

15. A process for producing a food comprising bCN and optionally also comprising gCN or its protein hydrolysates, the method comprising the following steps: I) Provide a heat-treated bCN concentrate according to one or more of claims 1-13 II) subjecting the heat-treated bCN concentrate to further processing, such as involving the hydrolysis of bCN and / or mixing with other components and / or filling the heat-treated bCN concentrate into a suitable container. Preferably, the food is infant formula, preferably containing an amount of bCN in the range of 1.0% w / w to 12% w / w relative to total solids, more preferably 1.3% w / w to 8% w / w relative to total solids, and most preferably in the range of 1.5% w / w to 6% w / w relative to total solids. Preferably, the food, more preferably infant formula, is produced by drying and blending powdered ingredients.

16. A food product obtainable by the process of claim 15, preferably, wherein the food product is an infant formula, wherein the food product preferably contains an amount of bCN in the range of 1.0% w / w to 12% w / w relative to total solids, more preferably 1.3% w / w to 8% w / w relative to total solids, and most preferably in the range of 1.5% w / w to 6% w / w relative to total solids. Preferably, the food, more preferably infant formula, is produced by drying and blending powdered ingredients.

17. Use of the heat-treated bCN concentrate according to any one or more of claims 1-13 as a food ingredient.

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